Engineered T cells. Function (growth factor / effector cytokine / effector molecules) Trafficking chemokine receptors
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1 Cell Processing Related Issues: Adoptive T cell Immunotherapy Bernard A. Fox Lab of Molecular and Tumor Immunology Robert W. Franz Cancer Research Center Providence Cancer Center Earle A. Chiles Research Institute Portland, Oregon, USA Molecular Microbiology and Immunology Tumor Immunology Focus Group OHSU Knight Cancer Institute Oregon Health and Science University Engineered T cells Marker Function (growth factor / effector cytokine / effector molecules) Trafficking chemokine receptors Resistance Gene Disruption: Steroid receptors (to provide resistance to steroids) TCR or targeting molecule 1
2 Characterization of procedure and product Source of cells cord blood Peripheral blood stem cells adult PBMC Tumor infiltrating lymphocytes (TIL) Variability of T cell donor Normal healthy donor Immunodefficient Patient Cancer patient Previous chemotherapy or radiation treatments Characterization of procedure and product Collection or recovery method Transport Conditions Apheresis: Cell selection device, density gradients, magnetic beads or FACS Surgery: TIL How many wells of tumor digest initiated (1, 6, 12, 24) Does the number of lymphocytes/ml change the frequency of detecting tumor specific T cells (cloids). Genetic modification and delivery plasmid vs. viral vs.? Optimal point to genetically modify ( Early? Late?) Scale up (cloning plates to 10L bags) Open vs closed culture methods 2
3 Characterization of procedure and product Generating / expanding tumor specific T cells Stimulation conditions REP Autologous REP Allogeneic (donor screening / eligibility) 21 CFR Part 1271 Eligibility Determination for Donors of Human Cells, Tissues and Cellular and Tissue Based Products (HCT/Ps) and Guidance for Industry: Eligibility Determination for Donors of Human Cells, Tissues, and Cellular and Tissue Based Products (HCT/Ps) Beads + Antibodies + Cytokine(s) APC + Antigen + Cytokine(s) + Costim (?) Autologous APC Artificial APC Characterization of procedure and product Maintaining cell phenotype in vivo replication engraftment potential / Memory cells killing capacity or cytokine release / Effector cells 3
4 Vendor source for material Medium / antibiotics FDA recommends that you do not use beta lactam antibiotics during the manufacturing of a therapeutic product Serum / human AB serum obtained from an approved blood bank and meets all blood donor criteria. Screening sera for desired growth support Growth factors /cytokines cgmp Recombinant available? Transduce feeder or stimulator cells to provide recombinant cytokine Enzymes Trypsin free of porcine parvovirus Triple enzymes : non cgmp (Sigma) vs cgmp Hyaluronidase, Hylenex, Baxter, or Amphadase, Amphastar Pharmaceuticals DNase I recombinant, grade I, Roche Liberase MTF C/T, Roche Monoclonal antibodies Points to Consider in the Manufacture and Testing of Monoclonal Antibody Products for Human Use Cell separation devices Plastics / availability Optimizing Logistics Culture monitoring metabolites, cell growth, cell cycle fraction.what is limiting to cell expansion? Central processing site(s) and qualification/comparability 4
5 Characterization of procedure and product Logistic Release/characterization assays; Sterility, Phenotype Potency Develop assay to assess functional property conferred by the genetic modification. Safety Carcinogenetic potential and how to assess Animal In vitro Suicide Vector 5
6 Earle A. Chiles Res. Inst. Christian Poehlein Shawn Jensen Tarsem Moudgil Michael LaCelle Kevin Friedman Chris Twitty James A. Thompson Ulf Petrausch Sachin Puri Levi Maston Sarah Church Edwin Walker Dan Haley Nelson Sanjaun Liliana Gonzalez Tanisha Snyder Andy Weinberg Hong-Ming Hu Lixin Wang Yuhan Li Walter Urba John Smith Brendan Curti Todd Crocenzi Rachel Sanborn Craig Nichols Lynn Glenn Brenda Fisher Theresa Ratzow Tracie Groupe Tracy Kelly Scot Larry Rachel Sautter Erin Sauter Julie Cramer LMU Dominik Ruettinger Hauke Winter Ilka Assmann Rudi Hatz Natasja van den Engel Dolores Schendel Heike Pohla Stephie Grützner Marcus Schlemmer Karl Walter Jauch Xi an Jun Ma L. Si Yili Wang Mayo Clinic AZ Helen Ross Janet Siebert 6
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